The cataract causing Cx50-S50P mutant inhibits Cx43 and intercellular communication in the lens epithelium.
The cataract causing Cx50-S50P mutant inhibits Cx43 and intercellular communication in the lens epithelium.
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DOI：
10.1016/j.yexcr.2009.01.017
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发表时间：
2009-04-01
影响因子：
3.7
通讯作者：
White TW
中科院分区：
文献类型：
--
作者：
DeRosa AM;Meşe G;Li L;Sellitto C;Brink PR;Gong X;White TW
连接蛋白50(CX50)的突变会导致人类和小鼠的白内障。连接蛋白突变导致多种白内障的机制(S)尚未完全阐明。在这里，我们测试了诱发白内障的CX50-S50P突变体是否与野生型连接蛋白43(Cx43)相互作用形成功能减弱的混合通道。通过双全细胞电压钳、免疫荧光显微镜和原位染料转移分析，我们发现突变亚基与野生型Cx43之间存在独特的相互作用。在非洲爪哇卵母细胞中，Cx50-S50P和Cx43的共表达使电偶联≥减少了90%，而蛋白表达没有减少。在转基因细胞中，CX50-S50P本身并不针对细胞-细胞界面，但CX50-S50P与Cx43的共表达导致其定位于细胞-细胞接触区。我们使用CX43条件性基因敲除、CX50基因敲除和CX50-S50P突变小鼠在体内检验了这种相互作用。在晶状体上皮细胞中同时表达Cx43和CX50-S50P的小鼠显示了Cx43的独特表达模式和Cx43蛋白的减少。原位染料转移实验表明，CX50-S50P突变体，而不是CX50，或CX43条件性敲除，以类似于CX43和CX50双重敲除的方式，极大地抑制了上皮细胞缝隙连接通讯。CX50-S50P的抑制作用导致体外电耦合减弱，以及上皮细胞染料渗透明显减少。这些结果提示，Cx43介导的上皮细胞偶联的显性抑制可能在CX50-S50P突变引起的晶状体病理生理过程中起作用。
Mutations in Connexin50 (Cx50) cause cataracts in both humans and mice. The mechanism(s) behind how mutated connexins lead to a variety of cataracts have yet to be fully elucidated. Here, we tested whether the cataract inducing Cx50-S50P mutant interacts with wild-type Connexin43 (Cx43) to form mixed channels with attenuated function. Using dual whole-cell voltage clamp, immunofluorescent microscopy and in situ dye transfer analysis we identified a unique interaction between the mutant subunit and wild-type Cx43. In paired Xenopus oocytes, co-expression of Cx50-S50P with Cx43 reduced electrical coupling ≥90%, without a reduction in protein expression. In transfected cells, Cx50-S50P did not target to cell-cell interfaces by itself, but co-expression of Cx50-S50P with Cx43 resulted in its localization at areas of cell-cell contact. We used Cx43 conditional knockout, Cx50 knockout and Cx50-S50P mutant mice to examine this interaction in vivo. Mice expressing both Cx43 and Cx50-S50P in the lens epithelium revealed a unique expression pattern for Cx43 and a reduction in Cx43 protein. In situ dye transfer experiments showed that the Cx50-S50P mutant, but not the Cx50, or Cx43 conditional knockout, greatly inhibited epithelial cell gap junctional communication in a manner similar to a double knockout of Cx43 and Cx50. The inhibitory affects of Cx50-S50P lead to diminished electrical coupling in vitro, as well as a discernable reduction in epithelial cell dye permeation. These data suggest that dominant inhibition of Cx43 mediated epithelial cell coupling may play a role in the lens pathophysiology caused by the Cx50-S50P mutation.
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影响因子：
64.5
作者：
Gong, XH;Li, E;Gilula, NB
通讯作者：
Gilula, NB
影响因子：
9.8
作者：
Mackay, D;Ionides, A;Bhattacharya, S
通讯作者：
Bhattacharya, S
影响因子：
4
作者：
Arora, A;Minogue, PJ;Berthoud, VM
通讯作者：
Berthoud, VM
影响因子：
9.8
作者：
Paznekas, WA;Boyadjiev, SA;Jabs, EW
通讯作者：
Jabs, EW
影响因子：
3.3
作者：
Du, X;Tabeta, K;Beutler, B
通讯作者：
Beutler, B